Structural organization of an intact phycobilisome and its association with photosystem II

Phycobilisomes (PBSs) are light-harvesting antennae that transfer energy to photosynthetic reaction centers in cyanobacteria and red algae. PBSs are supermolecular complexes composed of phycobiliproteins (PBPs) that bear chromophores for energy absorption and linker proteins. Although the structures of some individual components have been determined using crystallography, the three-dimensional structure of an entire PBS complex, which is critical for understanding the energy transfer mechanism, remains unknown. Here, we report the structures of an intact PBS and a PBS in complex with photosystem II (PSII) from Anabaena sp. strain PCC 7120 using single-particle electron microscopy in combination with biochemical and molecular analyses. In the PBS structure, all PBP trimers and the conserved linker protein domains were unambiguously located, and the global distribution of all chromophores was determined. We provide evidence that ApcE and ApcF are critical for the formation of a protrusion at the bottom of PBS, which plays an important role in mediating PBS interaction with PSII. Our results provide insights into the molecular architecture of an intact PBS at different assembly levels and provide the basis for understanding how the light energy absorbed by PBS is transferred to PSII.     

Structural modulation of gut microbiota during alleviation of type 2 diabetes with a Chinese herbal formula

The gut microbiota is hypothesized to have a critical role in metabolic diseases, including type 2 diabetes (T2D). A traditional Chinese herbal formula, Gegen Qinlian Decoction (GQD), can alleviate T2D. To find out whether GQD modulates the composition of the gut microbiota during T2D treatment, 187 T2D patients were randomly allocated to receive high (HD, n=44), moderate (MD, n=52), low dose GQD (LD, n=50) or the placebo (n=41) for 12 weeks in a double-blinded trial. Patients who received the HD or MD demonstrated significant reductions in adjusted mean changes from baseline of fasting blood glucose (FBG) and glycated hemoglobin (HbA1c) compared with the placebo and LD groups. Pyrosequencing of the V3 regions of 16S rRNA genes revealed a dose-dependent deviation of gut microbiota in response to GQD treatment. This deviation occurred before significant improvement of T2D symptoms was observed. Redundancy analysis identified 47 GQD-enriched species level phylotypes, 17 of which were negatively correlated with FBG and 9 with HbA1c. Real-time quantitative PCR confirmed that GQD significantly enriched Faecalibacterium prausnitzii, which was negatively correlated with FBG, HbA1c and 2-h postprandial blood glucose levels and positively correlated with homeostasis model assessment of β-cell function. Therefore, these data indicate that structural changes of gut microbiota are induced by Chinese herbal formula GQD. Specifically, GQD treatment may enrich the amounts of beneficial bacteria, such as Faecalibacterium spp. In conclusion, changes in the gut microbiota are associated with the anti-diabetic effects of GQD.     

Anaerobic ferrous oxidation by heterotrophic denitrifying enriched culture

Heterotrophic denitrifying enriched culture (DEC) from a lab-scale high-rate denitrifying reactor was discovered to perform nitrate-dependent anaerobic ferrous oxidation (NAFO). The DEC was systematically investigated to reveal their denitrification activity, their NAFO activity, and the predominant microbial population. The DEC was capable of heterotrophic denitrification with methanol as the electron donor, and autotrophic denitrification with ferrous salt as the electron donor named NAFO. The conversion ratios of ferrous-Fe and nitrate-N were 87.41 and 98.74 %, and the consumption Fe/N ratio was 2.3:1 (mol/mol). The maximum reaction velocity and half saturation constant of Fe were 412.54 mg/(l h) and 8,276.44 mg/l, and the counterparts of N were 20.87 mg/(l h) and 322.58 mg/l, respectively. The predominant bacteria were Hyphomicrobium, Thauera, and Flavobacterium, and the predominant archaea were Methanomethylovorans, Methanohalophilus, and Methanolobus. The discovery of NAFO by heterotrophic DEC is significant for the development of wastewater treatment and the biogeochemical iron cycle and nitrogen cycle.     

Mixed Polyethylene Glycol-Modified Breviscapine-Loaded Solid Lipid Nanoparticles for Improved Brain Bioavailability: Preparation,Characterization, and In Vivo Cerebral Microdialysis Evaluation in Adult Sprague Dawley Rats

Breviscapine is used in the treatment of ischemic cerebrovascular diseases, but it has a low bioavailability in the brain due to its poor physicochemical properties and the activity of P-glycoprotein efflux pumps located at the blood–brain barrier. In the present study, breviscapine-loaded solid lipid nanoparticles (SLN) coated with polyethylene glycol (PEG) derivatives were formulated and evaluated for their ability to enhance brain bioavailability. The SLNs were either coated with polyethylene glycol (40) (PEG-40) stearate alone (Bre-GBSLN-PS) or a mixture of PEG-40 stearate and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-PEG2000 (DSPE-PEG₂₀₀₀) (Bre-GBSLN-PS-DSPE) and were characterized both in vitro and in vivo. The mean particle size, polydispersity index, and entrapment efficiency for Bre-GBSLN-PS and Bre-GBSLN-PS-DSPE were 21.60 ± 0.10 and 22.60 ± 0.70 nm, 0.27 ± 0.01 and 0.26 ± 0.04, and 46.89 ± 0.73% and 47.62 ± 1.86%, respectively. The brain pharmacokinetic parameters revealed that the brain bioavailability of breviscapine from the Bre-GBSLN-PS and Bre-GBSLN-PS-DSPE was significantly enhanced (p < 0.01) with the area under concentration–time curve (AUC) of 1.59 ± 0.39 and 1.42 ± 0.58 μg h/mL of breviscapine, respectively, in comparison to 0.11 ± 0.02 μg h/mL from the commercial breviscapine injection. The ratios of the brain AUC for scutellarin in comparison with the plasma scutellarin AUC for commercial breviscapine injection, Bre-GBSLN-PS, and Bre-GBSLN-PS-DSPE were 0.66%, 2.82%, and 4.51%, respectively. These results showed that though both SLN formulations increased brain uptake of breviscapine, Bre-GBSLN-PS-DSPE which was coated with a binary combination of PEG-40 stearate and DSPE-PEG₂₀₀₀ had a better brain bioavailability than Bre-GBSLN-PS. Thus, the coating of SLNs with the appropriate PEG derivative combination could improve brain bioavailability of breviscapine and can be a promising tool for brain drug delivery.KEY WORDS: breviscapine, microdialysis, mixed PEGylation, P-glycoprotein (P-gp), solid lipid nanoparticles     

<Emphasis Type="Italic">fabC</Emphasis> of <Emphasis Type="Italic">Streptomyces lydicus</Emphasis> involvement in the biosynthesis of streptolydigin

Streptolydigin, a secondary metabolite produced by Streptomyces lydicus, is a potent inhibitor of bacterial RNA polymerases. It has been suggested that streptolydigin biosynthesis is associated with polyketide synthase (PKS) and nonribosomal peptide synthetase (NRPS). Thus, there is great interest in understanding the role of fatty acid biosynthesis in the biosynthesis of streptolydigin. In this paper, we cloned a type II fatty acid synthase (FAS II) gene cluster of fabDHCF from the genome of S. lydicus and constructed the SlyfabCF-disrupted mutant. Sequence analysis showed that SlyfabDHCF is 3.7 kb in length and encodes four separated proteins with conserved motifs and active residues, as shown in the FAS II of other bacteria. The SlyfabCF disruption inhibited streptolydigin biosynthesis and retarded mycelial growth, which were likely caused by the inhibition of fatty acid synthesis. Streptolydigin was not detected in the culture of the mutant strain by liquid chromatography–mass spectrometry. Meanwhile, the streptolol moiety of streptolydigin accumulated in cultures. As encoded by fabCF, acyl carrier protein (ACP) and β-ketoacyl-ACP synthase II are required for streptolydigin biosynthesis and likely involved in the step between PKS and NRPS. Our results provide the first genetic and metabolic evidence that SlyfabCF is shared by fatty acid synthesis and antibiotic streptolydigin synthesis.     

细菌在线虫生防中应用的研究进展

在具有杀线虫能力细菌的特征、作用方式及其在线虫生防中的潜在应用等方面,结合作者的研究工作,对国内、外的一些研究进展进行了综述,并提出了研究展望。     

Nitric Oxide Participates in the Induction of Brain Ischemic Tolerance via Activating ERK1/2 Signaling Pathways

The present study was undertaken to observe in vivo changes of expression and phosphorylation of ERK1/2 proteins during brain ischemic preconditioning and effects of inhibiting generation of nitric oxide (NO) on the changes to determine the role of ERKs in the involvement of NO participating in the acquired tolerance. Fifty-five Wistar rats were used. Brain ischemic preconditioning was performed with four-vessel occlusion for 3 min. Total ERK1/2 proteins and phospho-ERK1/2 in the CA1 hippocampus were assayed with Western immunoblot. Total ERK1/2 proteins did not change in period from 5 min to 5 days of reperfusion after preconditioning stimulus. While the level of phospho-ERK1/2 increased obviously to 223, 237, 300, 385 and 254% of sham level at times of 5 min, 2 h, 1, 3 and 5 days after preconditioning stimulus, respectively (P < 0.01). Administration of L-NAME, an inhibitor of NO synthase, 30 min prior to preconditioning stimulus failed to induce change in total ERK1/2 proteins (P > 0.05). However, phospho-ERK1/2 increased only to 138 and 176% of sham level at 2 h and 3 days after preconditioning stimulus, respectively, when animals were pretreated with L-NAME. The magnitudes of the increase were obviously low compared with those (237 and 385%) in animals untreated with L-NAME at corresponding time points (P < 0.01), which indicated that phosphorylation of ERK1/2 normally induced by preconditioning stimulus was blocked apparently by administration of L-NAME. The results suggested that phosphorylation of ERK1/2, rather than synthesis of ERK1/2 proteins, was promoted in brain ischemic preconditioning, and that the promotion was partly mediated by NO signal pathway.     

Molecular cloning and expression analyses of a new gene encoding 3-hydroxy-3-methylglutaryl-CoA synthase from Taxus × media

A new full-length cDNA encoding 3-hydroxy-3-methylglutaryl-CoA synthase (designated as TmHMGS, GenBank Accession No. AY644708), which catalyses the condensation of acetyl CoA and acetoacetyl CoA to form 3-hydroxy-3-methylglutaryl-CoA as an early step in the taxol biosynthetic pathway, was isolated from young leaves of Taxus × media by rapid amplification of cDNA ends (RACE) for the first time. The full-length cDNA of TmHMGS contained a 1431 bp open reading frame (ORF) encoding a deduced protein of 476 amino acid residues. The deduced protein had an isoelectric point of 5.23 and a calculated molecular mass of about 53 kDa. Amino acid sequence comparison analysis showed that TmHMGS had high similarity with a number of HMGSs ranging from Schizosaccharomyces pombe to humans, with much higher identity with other HMGSs from plants than those from yeast and humans. Phylogenic analysis showed that TmHMGS had closest relationship with HMGS from Pinus sylvestris. Tissue expression pattern analysis showed that TmHMGS expressed in needles and stems at similar level, but no expression could be detected in roots. Expression of TmHMGS was all induced by under different elicitors such as silver nitrate, ammonium ceric sulphate and methyl jasmonate, revealed that TmHMGS was an elicitor-responsive gene.     

Stage-specific expression of dynein light chain-1 and its interacting kinase, p21-activated kinase-1, in rodent testes: implications in spermiogenesis.

Mammalian spermatogenesis is a complex process involving regulatory interactions of many gene products. In this study, we found that dynein light chain-1 (DLC1), a component of the dynein motor complex, is highly expressed in mouse and rat testes. Immunohistochemically detectable levels of DLC1 are observed specifically in spermatids in steps 9-16 in distinct subcellular compartments: in steps 9-11, DLC1 is predominantly localized in the nucleus; in steps 12 and 13, it is found in both nucleus and cytoplasm; and in step 14-16, it is present exclusively in the cytoplasm. In addition, we found p21-activated kinase 1 (Pak1), a protein kinase that activates DLC1 by phosphorylating DLC1 at Serine 88, was also expressed during these stages of spermatogenesis. Pak1 was also expressed in Leydig cells, in preleptotene primary spermatocytes, and in round spermatids. The spermiogenic stage-specific expression of DLC1 suggests a role for DLC1 in chromatin condensation, spermatid shaping, and the final release of sperm from the spermatogenic epithelium. Further, Pak1 may also play a role in spermiogenesis by regulating DLC1 phosphorylation and, consequently, its function.     

Tests of candidate genes in breed cross populations for QTL mapping in livestock

In recent years, several F₂ crosses between outbred lines of livestock have been developed to identify quantitative trait loci (QTL). These populations are valuable for further genetic analysis, including positional candidate gene loci (CGL). Analysis of CGL in F₂ populations is, however, hindered by extensive between-breed linkage disequilibrium (LD). The objectives here were to develop and evaluate three tests for CGL in simulated F₂ breed-cross populations. 1) A standard association test, based on the fixed effect of CGL genotype. This test was significant for CGL at considerable distances from the QTL. 2) A marker-assisted association test, based on a test at the CGL of the fixed effect of CGL genotype in a breed-cross QTL interval mapping model. This removed the impact of between-breed LD, but was not powerful in detecting CGL closely linked to the QTL, unless the CGL was the QTL. 3) An F-drop test, comparing F ratios for a QTL at the CGL with and without the CGL included as fixed effect. It had low power to distinguish close from distant CGL. Power to distinguish two CGL within 10 cM from the QTL was limited and little improved by including QTL effects associated with markers to remove between-breed LD, although the power was greater when one of the CGL was the causative mutation. Therefore, while we conclude that candidate gene tests in QTL mapping populations must be interpreted with caution, we now have a clearer picture of the value of candidate gene tests in these populations.